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Serial analysis of ribosomal sequence tags (SARST): a high-throughput method for profiling complex microbial
Josh D Neufeld1, Zhongtang Yu, Wan Lam
1Department of Microbiology and Immunology, University of British Columbia, 6174 University Boulevard, Vancouver, British Columbia V6T 1Z3, Canada.
Environmental Microbiology
|February 6, 2004
Summary
Serial Analysis of Ribosomal Sequence Tags (SARST) offers a high-throughput method for profiling microbial diversity. This novel tool accurately reflects microbial community composition in soils and sediments for various applications.
Area of Science:
- Microbiology
- Molecular Biology
- Bioinformatics
Background:
- Microbial communities harbor vast phylogenetic diversity, necessitating advanced molecular tools for comprehensive analysis.
- Existing methods for studying microbial diversity, such as SSU rDNA clone libraries, have limitations in throughput.
Purpose of the Study:
- To introduce and validate Serial Analysis of Ribosomal Sequence Tags (SARST) as an innovative molecular tool.
- To assess the throughput and reproducibility of SARST for microbial diversity profiling.
Main Methods:
- SARST involves enzymatic amplification and ligation of ribosomal sequence tags (RSTs) from bacterial SSU rDNA V1-regions into concatemers.
- Concatemers are cloned and sequenced, yielding up to 20 RSTs per reaction, significantly increasing throughput.
- SARST performance was evaluated using defined mixtures of pure cultures and duplicate arctic soil DNA samples.
Main Results:
- SARST accurately reflected the composition of a defined microbial mixture.
- Duplicate soil library analyses demonstrated high correlation in RST profiles (r² = 0.80) and division-level distribution (r² = 0.99).
- Specific primers designed from abundant soil RSTs enabled successful amplification of larger SSU rDNA fragments for phylogenetic analysis.
Conclusions:
- SARST is a powerful, reproducible, and high-throughput method for microbial diversity profiling.
- The approach is suitable for applications in medical, industrial, and environmental microbiology.
- SARST facilitates deeper insights into the functional implications of microbial community structure.